Phenotyping of tumor infiltrating immune cells using mass-cytometry (CyTOF)

Abstract

The anti-tumor immune response plays a crucial role in cancer patient outcome as well as in response to the growing family of immunotherapeutic treatments. Improving patient prognostic and therapeutic management requires a better knowledge of the tumor microenvironment, for which a deep characterization of tumor-infiltrating immune populations is instrumental. Mass cytometry represents an excellent tool in tumor Immunology, as it allows the simultaneous analysis of >40 markers on single cells. In this chapter, we review challenging technical aspects of the mass cytometry phenotyping of tumor infiltrating immune cells, focusing on fresh human solid tumor samples. We first explain how to design mass cytometry experiments, then provide detailed protocols to isolate mononuclear immune cells from solid tissues and to stain them for an acquisition on a mass cytometer. We also discuss how to optimize the preparation of single immune cell samples, and how to ensure the reproducibility of data generated from distinct fresh human samples. Finally, we provide troubleshooting suggestions for difficult sample acquisitions on a mass cytometer.

Keywords: Cell isolation; CyTOF; Immune contexture; Inter-assay reproducibility; Mass cytometry; Panel design; Tumor infiltrating immune cells; Tumor infiltrating lymphocytes (TILs); Tumor microenvironment (TME).